Characterization of toxins of Clostridia causing gas-gangrene

Propagation of freeze dried ATCC cultures of Gas-gangrene Clostridium perfringens, Clostridium septicum and Clostridium novyi was done in Reinforced Clostridial Medium (ReCM). The propagated culture was inoculated in Robertson's Cooked Meat Medium (RCMM) for toxin production in bulk amount. Lethality assay of the crude toxin obtained was checked in mice. Histopathological studies of the liver, heart, lung and kidney indicated the damage of blood vascular system, necrosis of vascular endothelium and hemorrhages. Further purification of the crude toxin was done by DEAE-cellulose chromatography and Sephadex G-100 column chromatography. The LD50 of the purified toxin was worked out. The LD50 of C.perfringens, C.septicum and C.novyi was 10^4.1LD50/ml, 10^3.2LD50/ml and 10^2.9LD50/ml. The molecular weight of the protein fraction of purified toxin of C.perfringens, C.septicum and C.novyi were 30KD, 46KD and 200KD respectively.